The use of SNPs as polymorphic markers for linkage disequilibrium (LD) studies offers great potential. Their lack of informativeness is believed to be mostly compensated by 1) their number, over 1.4 million, and 2) their relatively homogeneous distribution throughout the human genome. In certain embodiments for genes containing multiple SNPs, haplotype structure rather than individual SNPs can be the principal determinant of phenotypic consequences. In addition, haplotype analysis is a popular tool in assessing LD and for mapping complex disease genes in association studies where SNP-phenotype associations may not be detectable. Numerous studies have indicated that haplotypes can provide additional power in detecting association.
Haplotypes are the collection of genotypes that occur in a single allele or chromosome. Due to the diploidy nature of the human genome, the challenge is to obtain genotype data from a single chromosome or allele. There are two ways to perform haplotyping: 1) Computational haplotyping and 2) Molecular haplotyping. The first involves converting diploidic genotype information into estimated haplotypes. This method offers only a statistical approximation of an individual's haplotype structure. The second method provides haplotype determination through physical allele selection followed by genotyping analysis. Such a molecular approach may provide a more accurate picture.
No high-throughput methods for allele selection are available. One available method for obtaining a specific allele involves converting diploidic cells to haploidic clones. This, however, is a cost and time prohibitive process. Other approaches use conventional Allele-Specific-PCR/genotyping. A challenge with conventional Allele-Specific PCR (ASPCR) is potential for amplification leakage, i.e., amplification of the undesired wrong allele. Accordingly, there is a need for methods for sequence analysis of specific alleles that reduces allele leakage, and methods that permit high-throughput analysis. Accordingly, among the objects herein, it is an object to provide such methods.